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Acta Agronomica Sinica ›› 2022, Vol. 48 ›› Issue (4): 801-811.doi: 10.3724/SP.J.1006.2022.14077

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Genome editing of BnMLO6 gene by CRISPR/Cas9 for the improvement of disease resistance in Brassica napus L

SHI Yu-Qin1,2(), SUN Meng-Dan1, CHEN Fan1, CHENG Hong-Tao1,2, HU Xue-Zhi1, FU Li1, HU Qiong1,2, MEI De-Sheng1,2,*(), LI Chao1,*()   

  1. 1Oil Crops Research Institute, Chinese Academy of Agricultural Sciences / Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture and Rural Affairs, Wuhan 430062, Hubei, China
    2Graduate School of Chinese Academy of Agricultural Sciences, Beijing 100081, China
  • Received:2021-04-25 Accepted:2021-07-12 Online:2022-04-12 Published:2021-08-13
  • Contact: MEI De-Sheng,LI Chao E-mail:shiyuqin514@163.com;meidesheng@caas.com;lichao01@caas.cn
  • Supported by:
    Agricultural Science and Technology Innovation Program of Chinese Academy of Agricultural Sciences

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Abstract:

Gene editing technology can modify the target gene efficiently and accurately, which opens up a new way for crop genetic improvement. Mildew resistance locus O (MLO) gene is a key negative regulator of plant defense against powdery mildew. Mutation of MLO gene can enhance plant resistance to powdery mildew, but whether it has the same function is not reported in oilseed rape. In this study, the relative expression analysis suggested that BnMLO6 gene was induced by Sclerotinia sclerotiorum. To explore the potential role of BnMLO6 gene in pathogen resistance, six homologous copies of BnMLO6 gene mutated synchronously by CRISPR/Cas9 gene editing technology and mlo6-212 mutant line was generated for further analysis. Genetic analysis revealed that CRISPR/Cas9 induced mutagenesis of BnMLO6 gene could be stably inherited. In addition, mlo6-212 mutant line indicated obvious resistance to powdery mildew in both field and greenhouse condition. The lesion area of mlo6-212 mutant was reduced by 19.5% after 24 hours inoculation with S. sclerotiorum. Meanwhile, mutation of BnMLO6 gene could stimulate the spontaneous accumulation of callose in leaves and activate ethylene and jasmonic acid transduction pathway. Thus, BnMLO6 gene was probably involved in multiple pathogen resistance pathways to negatively regulate resistance to powdery mildew and S. sclerotiorum in oilseed rape. The results not only provide theoretical basis for the study of BnMLO6 involved resistance regulation of multiple pathogens, but also provide resistant resources and technical support for genetic improvement of disease resistance in oilseed rape.

Key words: oilseed rape, genome editing, MLO, powdery mildew, Sclerotinia sclerotiorum

Table 1

List of PCR primers"

引物名称
Primer name		 引物序列
Primer sequence (5¢-3¢)		 扩增产物
Product length (bp)
Bnactin-RT-F TCTGGCATCACACTTTCTACAACGAGC 688
Bnactin-RT-R CAGGGAACATGGTCGAACCACC
MLO6-A03-RT-F2 ATCAGAAGAGTTGCAAGAGTATCC 603
MLO6-A03-RT-R2 GCCACAAACCAGATCACAGGAC
MLO6-C03-RT-F2 GCAGTAAATCCGAAGAGTTGAAAG 617
MLO6-C03-RT-R3 TGGTTGTGAGGAGGAATAGCAC
MLO6-A01-RT-F1 AGAACAAAAAAGCACTGTATGAAGG 1096
MLO6-A01-RT-R1 CACAGTTCTTGAGCTTGAACTCATA
MLO6-C01-RT-R1 TTCTTGTGGAAGCAGTTCTTGAG 952
MLO6-C01-RT-F2 CTCCGAGAGCATTGCAGCATCA
MLO6-A09-RT-F2 AGTGTGCAGAGAAGGGAAAAGTC 846
MLO6-A09-RT-R2 AGAGTTTCTTGGAGACAGCGTGT
MLO6-C09-RT-R1 TGTGTGTGGAACTTACCTGAGTCA 835
MLO6-C09-RT-F2 TGCAGAGAAGGGAAAGGTTGCT
NPTII-F GATGGATTGCACGCAGGT 1050
NPTII-R TCGTCAAGAAGGCGATAGA
MLO6-A03-F2 ATTTGGGTTTTTGTTACATATACCAATGTATA 669
MLO6-A03-R2 CTCTTTGTGTGTATGTGTTTGTGACTTACC
MLO6-C03-F2 TTCGGTTTTTTTTTTTTTTACATGAACC 678
MLO6-C03-R2 GAAAGATGTTAGTTTGATGTTTTCTCTCTG
MLO6-A01-F1 GGTCTTAACGGCATAGCAAAACGAT 677
MLO6-A01-R2 CGTGGAATTATTTTGTTTGCTTACCGT
MLO6-C01-F1 TTCTTACCGCATTGCAAAATGATTCTT 680
MLO6-C01-R2 TGTGGGATACTACTTTGCTTGCTTACC
MLO6-A09-F1 CTCAGAATGTTTCTGCATCCGATATCT 510
MLO6-A09-R3 CTTTTACTTACCCAACTGAAATTCTGATGAC
MLO6-C09-R1 GAATTGCTCTGCTTGCTTACCGTTG 619
MLO6-C09-F2 CAACAATCCCAGAACTAATATATTTGCTCT

Table 2

Primers of disease resistance related genes"

基因名称
Gene name		 油菜中基因号
Gene name of B. napus		 引物名称
Primer name		 引物序列
Primer sequence (5¢-3¢)		 扩增产物
Product length (bp)
BnORA059 BnaA10G0042700ZS ORA59-2-F ATCAATCCTTCCTCTCAGTTAGC 517
ORA59-2-R TCTTGATCCGTAACAATACTCTG
BnaC05G0044000ZS ORA59-3-F CAATCATTCCTCTCAGCTGTTAGC 553
ORA59-3-R CTTGATTCGTAACAACACTTTGTT
BnPR4 BnaA03G0296000ZS PR4-1-F CGCGTTTGCGGCTAAAACA 130
PR4-1-R TGGTTTCCTTTCCACGTTGAG
BnaA03G0296200ZS PR4-3-F CAGACTTAGCATAGCCATCATAT 214
PR4-3-R GCTCTCTCCAGAGCCGCTT
BnaC03G0354600ZS PR4-5-F AGCCGCTCAATCCGCTAATA 194
PR4-5-R TACCGCAAGAATCACGACCA
基因名称
Gene name		 油菜中基因号
Gene name of B. napus		 引物名称
Primer name		 引物序列
Primer sequence (5¢-3¢)		 扩增产物
Product length (bp)
BnPR4 BnaC03G0354700ZS PR4-6-F CAGACTTAGCATAACCCTCGTA 213
PR4-6-R CTCTCTCCGGAACCGCTTG
BnERF1 BnaA01G0293300ZS ERF1-1-F TCGTCCAACAACTACTCTCTCC 147
ERF1-1-R GTACGACTTCTCTGGCTTTCTTG
BnaC01G0362300ZS ERF1-2-F GGAATCTTTCTCCTCCTCGTCTT 153
ERF1-2-R TGACTTCTCTGGTTTTCTCGATG
BnERF2 BnaA09G0212800ZS ERF2-3-F AGATTTAGAACCGTCGTCGAAG 409
ERF2-3-R TATTTCTCCTCCGTTTCGGTGT
BnaC02G0416900ZS ERF2-4-F GAGTTGTATCACCGAATTTGTAG 320
ERF2-4-R ACGAAGACGATGAAGACGAAGTA
BnaC09G0247600ZS ERF2-6-F TAGATAAGCCCCCGGCTAATC 396
ERF2-6-R TCCGTTTCGGCGTCCCGTTA

Fig. 1

Induced expression of BnMLO6 genes by S. sclerotiorum"

Fig. 2

Design of sgRNA and construction of vector"

Fig. 3

PAGE-based genotyping of the BnMLO6-C01 T0 generation plants M: DNA marker; the triangle arrow represents the edit strip."

Fig. 4

Mutation of targeted sites of MLO homologous in mlo6-212 T0 plants A: mutation of DNA sequence, the underlined parts represent PAM sites; B: change of amino acid sequence, and the red parts indicate mutations."

Table 3

Partial editing results of T1 generation of mlo6-212"

T1代编号
Number of T1 generation		 MLO6-A03 MLO6-C03 MLO6-A01 MLO6-C01 MLO6-A09 MLO6-C09
mlo6-212-1 -15 bp -13 bp +1 bp -12 bp -3 bp -2 bp
mlo6-212-3 -15 bp -13 bp -2 bp 无No mutation -3 bp -2 bp
mlo6-212-4 -15 bp -2 bp -2 bp -12 bp -3 bp -2 bp
mlo6-212-7 -15 bp -13 bp +1 bp 无No mutation 无No mutation 无No mutation
mlo6-212-8 -15 bp -13 bp -2 bp -12 bp -3 bp -2 bp
mlo6-212-9 -15 bp -13 bp -2 bp -12 bp -3 bp -2 bp
mlo6-212-10 -15 bp -13 bp +1 bp -12 bp 无No mutation 无 No mutation

Fig. 5

Infection of powdery mildew in T1 generation A: field powdery mildew infection; B: indoor powdery mildew infection, bar: 1 cm; C: field susceptibility of mlo6-212 lines to powdery mildew in T2 generation."

Fig. 6

Microscopic observation and analysis of callose in T1 generation A: microscopic observation of callose, the black arrow indicates callose, bar: 30 mm; B: the number of callose deposits per field area."

Fig. 7

Assessment of stem rot resistance by inoculation of S. sclerotiorum in T2 generation A: 24 hours after S. sclerotiorum infection; B: 36 hours after S. sclerotiorum infection; bars: 1 cm; C: analysis of lesion area. *: significant difference at P < 0.05 between the two groups."

Fig. 8

Expression patterns of JA/ET signaling pathway genes before and after inoculation Each row represents the expression change of a homologous copy of the related gene; from top to bottom, the three copies of BnERF2 are BnaA09G0212800ZS, BnaC02G0416900ZS, and BnaC09G0247600ZS; the two copies of BnERF1 are BnaA01G0293300ZS, and BnaC01G0362300ZS; the four copies of BnPR4 are BnaA03G0296000ZS, BnaA03G0296200ZS, BnaC03G0354600ZS, and BnaC03G0354700ZS; the two copies of BnOAR059 are BnaA10G0042700ZS and BnaC05G0044000ZS; samples of the order from left to right is wild-type uninfected, mutant infected, wild-type infected 24 hours, mutant infected 24 hours, wild-type infected 36 hours, and mutant uninfected 36 hours."

[1] 李利霞, 陈碧云, 闫贵欣, 高桂珍, 许鲲, 谢婷, 张付贵, 伍晓明. 中国油菜种质资源研究利用策略与进展. 植物遗传资源学报, 2020, 21:1-19.
Li L X, Chen B Y, Yan G X, Gao G Z, Xu K, Xie T, Zhang F G, Wu X M. Proposed strategies and current progress of research and utilization of oilseed rape germplasm in China. J Plant Genet Resour, 2020, 21:1-19 (in Chinese with English abstract).
[2] 刘成, 冯中朝, 肖唐华, 马晓敏, 周广生, 黄凤洪, 李加纳, 王汉中. 我国油菜产业发展现状、潜力及对策. 中国油料作物学报, 2019, 41:485-489.
Liu C, Feng Z C, Xiao T H, Ma X M, Zhou G S, Huang F H, Li J N, Wang H Z. Development, potential and adaptation of Chinese rapeseed industry. Chin J Oil Crop Sci, 2019, 41:485-489 (in Chinese with English abstract).
[3] 孙祥良, 王华弟, 曹奎荣, 朱金良. 油菜菌核病对油菜千粒重及产量的影响. 浙江农业科学, 2014, 11:1732-1733.
Sun X L, Wang H D, Cao K R, Zhu J L. Effects of Sclerotinia sclerotinia on 1000-grain weight and yield of rapeseed. J Zhe jiang Agric Sci, 2014, 11:1732-1733 (in Chinese with English abstract).
[4] 吴健, 周永明, 王幼平. 油菜与核盘菌互作分子机理研究进展. 中国油料作物学报, 2018, 40:721-729.
Wu J, Zhou Y M, Wang Y P. Research progress on molecular mechanisms of Brassica napus-Sclerotinia sclerotiorum interaction. Chin J Oil Crop Sci, 2018, 40:721-729 (in Chinese with English abstract).
[5] 杨清坡, 刘万才, 黄冲. 近10年油菜主要病虫害发生危害情况的统计和分析. 植物保护, 2018, 44(3):24-30.
Yang Q P, Liu W C, Huang C. Statistics and analysis of oilseed rape losses caused by main diseases and insect pests in recent 10 years. Plant Prot, 2018, 44(3):24-30 (in Chinese with English abstract).
[6] Gaetán S, Madia M. First report of canola powdery mildew caused by Erysiphe polygoni in Argentina. Plant Dis, 2004, 88:1163.
doi: 10.1094/PDIS.2004.88.10.1163C pmid: 30795271
[7] 邵登魁. 油菜抗白粉病鉴定及相关的生理生化特性研究. 甘肃农业大学硕士学位论文,甘肃兰州, 2006.
Shao D K. Identification of Resistance to Erysiphe cruciferarum Junell and Study on Enzymes Associated with PM in Brassica Rape. MS Thesis of Gansu Agricultural University, Lanzhou, Gansu,China, 2006 (in Chinese with English abstract).
[8] Tyagi S, Kumar R, Kumar V, Won S Y, Shukla P. Engineering disease resistant plants through CRISPR-Cas9 technology. GM Crop Food, 2021, 12:125-144.
[9] 单奇伟, 高彩霞. 植物基因组编辑及衍生技术最新研究进展. 遗传, 2015, 37:953-973.
Shan Q W, Gao C X. Research progress of genome editing and derivative technologies in plants. Hereditas, 2015, 37:953-973 (in Chinese with English abstract).
[10] 刘耀光, 李构思, 张雅玲, 陈乐天. CRISPR/Cas植物基因组编辑技术研究进展. 华南农业大学学报, 2019, 40(5):38-49.
Liu Y G, Li G S, Zhang Y L, Chen L T. Current advances on CRISPR/Cas genome editing technologies in plants. J South China Agric Univ, 2019, 40(5):38-49 (in Chinese with English abstract).
[11] Cong L, Ran F A, Cox D, Lin S, Barretto R, Habib N, Hsu P D, Wu X, Jiang W, Marraffini L A, Zhang F. Multiplex genome engineering using CRISPR/Cas systems. Science, 2013, 339:819-823.
doi: 10.1126/science.1231143 pmid: 23287718
[12] Sorek R, Lawrence C M, Wiedenheft B. CRISPR-mediated adaptive immune systems in bacteria and archaea. Annu Rev Biochem, 2013, 82:237-266.
doi: 10.1146/biochem.2013.82.issue-1
[13] Mao Y, Zhang H, Xu N, Zhang B, Gou F, Zhu J K. Application of the CRISPR-Cas system for efficient genome engineering in plants. Mol Plant, 2013, 6:2008-2011.
doi: 10.1093/mp/sst121
[14] Xie K, Yang Y. RNA-guided genome editing in plants using a CRISPR-Cas system. Mol Plant, 2013, 6:1975-1983.
doi: 10.1093/mp/sst119
[15] Upadhyay S K, Kumar J, Alok A, Tuli R. RNA-guided genome editing for target gene mutations in wheat. G3: Gen Genom Genet, 2013, 3:2233-2238.
[16] Liang Z, Zhang K, Chen K, Gao C. Targeted mutagenesis in Zea mays using TALENs and the CRISPR/Cas system. J Genet Genomics, 2014, 41:63-68.
doi: 10.1016/j.jgg.2013.12.001 pmid: 24576457
[17] Gao J, Wang G, Ma S, Xie X, Wu X, Zhang X, Wu Y, Zhao P, Xia Q. CRISPR/Cas9-mediated targeted mutagenesis in Nicotiana tabacum. Plant Mol Biol, 2015, 87:99-110.
doi: 10.1007/s11103-014-0263-0
[18] Soyk S, Müller N A, Park S J, Schmalenbach I, Jiang K, Hayama R, Zhang L, Van Eck J, Jiménez-Gómez J M, Lippman Z B. Variation in the flowering gene SELF PRUNING 5G promotes day-neutrality and early yield in tomato. Nat Genet, 2017, 49:162-168.
doi: 10.1038/ng.3733
[19] Braatz J, Harloff H J, Mascher M, Stein N, Himmelbach A, Jung C. CRISPR-Cas9 targeted mutagenesis leads to simultaneous modification of different homoeologous gene copies in polyploid oilseed rape (Brassica napus). Plant Physiol, 2017, 174:935-942.
doi: 10.1104/pp.17.00426
[20] Jørgensen I H. Discovery, characterization and exploitation of MLO powdery mildew resistance in barley. Euphytica, 1992, 63:141-152.
doi: 10.1007/BF00023919
[21] Devoto A, Hartmann H A, Piffanelli P, Elliott C, Simmons C, Taramino G, Goh C S, Cohen F E, Emerson B C, Schulze-Lefert P, Panstruga R. Molecular phylogeny and evolution of the plant- specific seven-transmembrane MLO family. J Mol Evol, 2003, 56:77-88.
doi: 10.1007/s00239-002-2382-5
[22] Liu Q, Zhu H. Molecular evolution of the MLO gene family in Oryza sativa and their functional divergence. Gene, 2008, 409:1-10.
doi: 10.1016/j.gene.2007.10.031
[23] Konishi S, Sasakuma T, Sasanuma T. Identification of novel MLO family members in wheat and their genetic characterization. Genes Genet Syst, 2010, 85:167-175.
pmid: 21041976
[24] Zhou S J, Jing Z, Shi J L. Genome-wide identification, characterization, and expression analysis of the MLO gene family in Cucumis sativus. Genet Mol Res, 2013, 12:6565-6578.
doi: 10.4238/2013.December.11.8 pmid: 24391003
[25] Wolter M, Hollricher K, Salamini F, Schulze-Lefert P. The mlo resistance alleles to powdery mildew infection in barley trigger a developmentally controlled defence mimic phenotype. Mol Gen Genet, 1993, 239:122-128.
doi: 10.1007/BF00281610
[26] Ropenack E V, Parr A, Schulze-Lefert P. Structural analyses and dynamics of soluble and cell wall-bound phenolics in a broad spectrum resistance to the powdery mildew fungus in barley. J Biol Chem, 1998, 273:9013-9022.
doi: 10.1074/jbc.273.15.9013
[27] Piffanelli P, Zhou F, Casais C, Orme J, Jarosch B, Schaffrath U, Collins N C, Panstruga R, Schulze-Lefert P. The barley MLO modulator of defense and cell death is responsive to biotic and abiotic stress stimuli. Plant Physiol, 2002, 129:1076-1085.
pmid: 12114562
[28] Kuhn H, Lorek J, Kwaaitaal M, Becker K, Micali C, Ver Loren van Themaat E, Bednarek P, Raaymakers T M, Appiano M, Bai Y, Meldau D, Baum S, Conrath U, Feussner I, Panstruga R. Key components of different plant defense pathways are dispensable for powdery mildew resistance of the Arabidopsis mlo2 mlo6 mlo12 triple mutant. Front Plant Sci, 2017, 8:1006.
doi: 10.3389/fpls.2017.01006
[29] 贾云飞, 张国海, 刘崇怀, 樊秀彩, 姜建福, 孙海生, 张颖. Mlo基因在葡萄抗白腐病中作用的研究. 植物生理学报, 2017, 53:1649-1658.
Jia Y F, Zhang G H, Liu C H, Fan X C, Jiang J F, Sun H S, Zhang Y. Study on the function of resistance to white rot of Mlo genes in grapevine. Plant Physiol J, 2017, 53:1649-1658 (in Chinese with English abstract).
[30] Acevedo-Garcia J, Gruner K, Reinstädler A, Kemen A, Kemen E, Cao L, Takken F L W, Reitz M U, Schäfer P, O’Connell R J, Kusch S, Kuhn H, Panstruga R. The powdery mildew-resistant Arabidopsis mlo2 mlo6 mlo12 triple mutant displays altered infection phenotypes with diverse types of phytopathogens. Sci Rep, 2017, 7:9319.
doi: 10.1038/s41598-017-07188-7 pmid: 28839137
[31] McGrann G R, Stavrinides A, Russell J, Corbitt M M, Booth A, Chartrain L, Thomas W T, Brown J K. A trade off between mlo resistance to powdery mildew and increased susceptibility of barley to a newly important disease, Ramularia leaf spot. J Exp Bot, 2014, 65:1025-1037.
doi: 10.1093/jxb/ert452 pmid: 24399175
[32] Shi J, Wan H, Zai W, Xiong Z, Wu W. Phylogenetic relationship of plant MLO genes and transcriptional response of MLO genes to Ralstonia solanacearum in tomato. Genes, 2020, 11:487.
doi: 10.3390/genes11050487
[33] Ma X, Zhang Q, Zhu Q, Liu W, Chen Y, Qiu R, Wang B, Yang Z, Li H, Lin Y, Xie Y, Shen R, Chen S, Wang Z, Chen Y, Guo J, Chen L, Zhao X, Dong Z, Liu Y G. A robust CRISPR/Cas9 system for convenient, high-efficiency multiplex genome editing in monocot and dicot plants. Mol Plant, 2015, 8:1274-1284.
doi: 10.1016/j.molp.2015.04.007
[34] Li C, Hao M, Wang W, Wang H, Chen F, Chu W, Zhang B, Mei D, Cheng H, Hu Q. An efficient CRISPR/Cas9 platform for rapidly generating simultaneous mutagenesis of multiple gene homeologs in allotetraploid oilseed rape. Front Plant Sci, 2018, 9:442.
doi: 10.3389/fpls.2018.00442
[35] Luna E, Pastor V, Robert J, Flors V, Mauch-Mani B, Ton J. Callose deposition: a multifaceted plant defense response. Mol Plant Microbe Interact, 2011, 24:183-193.
doi: 10.1094/MPMI-07-10-0149
[36] 何烈干, 宋来强, 汤洁, 周银生, 马辉刚. 油菜菌核病抗性鉴定方法比较及抗病种质资源的筛选. 江苏农业科学, 2018, 46(18):90-93.
He L G, Song L Q, Tang J, Zhou Y S, Ma H G. Comparison of identification methods for resistance to Sclerotinia sclerotiorum and screening of resistant materials of rapeseed. Jiangsu Agric Sci, 2018, 46(18):90-93 (in Chinese).
[37] Zaidi S S, Mukhtar M S, Mansoor S. Genome editing: targeting susceptibility genes for plant disease resistance. Trends Biotechnol, 2018, 36:898-906.
doi: 10.1016/j.tibtech.2018.04.005
[38] Sun Q, Lin L, Liu D, Wu D, Fang Y, Wu J, Wang Y. CRISPR/Cas9-mediated multiplex genome editing of the BnWRKY11 and BnWRKY70 genes in Brassica napus L. Int J Mol Sci, 2018, 19:2716.
doi: 10.3390/ijms19092716
[39] Naumann M, Somerville S, Voigt C. Differences in early callose deposition during adapted and non-adapted powdery mildew infection of resistant Arabidopsis lines. Plant Signal Behav, 2013, 8:e24408.
[40] Bari R, Jones J D G. Role of plant hormones in plant defence responses. Plant Mol Biol, 2009, 69:473-488.
doi: 10.1007/s11103-008-9435-0
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